1. Field of the Invention
The present invention relates to the regulation of cervical dilatation and extensibility by the use of nitric oxide donors and/or substrates or nitric oxide inhibitors.
2. Description of the Related Art
Parturition (expulsion of the fetus from the uterus), requires both contractions of the myometrium, the smooth muscle of the uterus, and a softening of the connective tissue of the cervix, so that it will stretch and dilate sufficiently to allow the fetus to be expelled. This softening is known as "ripening".
The current preferred method of cervical ripening is by the use of prostaglandin B.sub.2. This is used as a vaginal gel or tablet or as a gel placed in the cervix. One worry about the use of prostaglandin E.sub.2 is that there is a possibility of hyperstimulation of the uterus, leading to excessively strong myometrial contractions before the cervix is ripened and therefore before a comfortable or safe birth is possible.
The ideal preparation would soften and efface the cervix without causing myometrial contractions. This would allow the subsequent contractions (induceable if necessary with a small dose of prostaglandin) to deliver the baby with a minimum of resistance. There is good evidence from animal experiments that the antiprogestins such as RU486 would meet these requirements, but the problem with this drug is that it has associated antiglucocorticoid activity which might be detrimental to the fetus.
One of the most exciting recent advances in biology and medicine is the discovery that nitric oxide is produced by endothelial cells and that it is involved in the regulation of vascular tone, platelet aggregation, neurotransmission and immune activation (Furchgott and Zawadzki, 1980; Moncada, Palmer and Higgs, 1991; Ignarro, 1991). Nitric oxide is an important mediator of relaxation of the muscular smooth muscle (Montada, Palmer and Higgs, 1991) and was formerly known as EDRF (endothelin-derived relaxing factor) (Furchgott und Zawadzki, 1980; Moncada, Palmer and Higgs, 1991).
Nitric oxide is synthesized by the oxidative deamination of a guanidino nitrogen of L-arginine by at least different isoforms of a flavin-containing enzyme, nitric oxide synthase (Montada, Palmer and Higgs, 1991).
Nitric oxide can also be generated by application of various nitric oxide donors such as sodium nitroprusside, nitroglycerin, glyceryl trinitrate, SIN-1, isosorbid mononitrate, isosorbid di nitrate, etc.
Synthesis of nitric oxide has been shown to be competitively inhibited by analogies of L-arginine; NG-nitro-Larginine methyl ester (L-NAME), NG-monoethyl-L-arginine (LMMA), N-iminoethyl-L-amithine (L-NIO), L-monomethyl-Larginine (L-NNMA), L-NG-methylarginine (LNMA), Nw-nitro-L-arginine (L-NA) and Aminaguanidine. Treatment of nonpregnant guinea pigs with L-NAME results in increased uterine contractility. Thus, inhibition of nitric oxide synthase-stimulated uterine contractility indicates that the tonic release of nitric oxide maintains the uterus in a quiescent state. Similarly, treatment of pregnant guinea pigs with L-NAME induced preterm labor. On the other hand, treatment of rat uterine strips in vitro with L-arginine inhibited contractions. These studies show that nitric oxide production by the uterus inhibits uterine contractility and a blockade of this synthesis results in increased muscle contractility both in pregnant and nonpregnant animals. Thus, nitric oxide substrates or donors are useful therapeutically to prevent uterine contractility and nitric oxide inhibitors are effective in stimulating uterine contractions.
Based on preliminary studies using rhesus monkees, Jennings et al. (The Journal of Maternal-Fetal Medicine 2:170-175 (1993)) have suggested that nitric oxide may be important in uterine quisence during pregnancy, and exogenous nitric oxide may be useful in controlling preterm labour.
Nitric oxide synthase (NOS) activity was demonstrated in multiple structures within the gravid rat uterus by Natuzzi et al. (Biochem. and Biophys. Res. Commun., 194, No.1, 1-8, (1993)) and they conclude NOS to be present within multiple structures of the gravid rat uterus. Reduction in NOS activity at parturition suggests NO may contribute to the maintenance of uterine contractile quisence during gestation.
Also, morphological studies have already shown the occurence of NOS (NADPH-diaphorase, a histochemical method detecting all NOS isoforms) and NO metabolizing enzymes such as superoxide dismutase in the cervix (Papka et al., Neuroscience Letters 147 (1992); Shiotani et al., Acta Histochem. Cytochem. 26:57-64 (1993)).
It has now been found that L-NAME treatment s.c. in pregnant guinea pigs on days 48-49 p.c. (term day 67+3 days) surprisingly led to a reduction in the cervical extensibility and a reduction in the cervical dilatation.
In pregnant rats, an increased production of nitric oxide (parameter, nitrite and nitrate release in vitro) from the uterine cervix has been found at the time of delivery in comparison with day 18 of pregnancy. In contrast there was a reduction in the uterine (myometrial) NO production in the course of rat pregnancy. The antiprogestins onapristone increased the nitric oxide production in the uterine cervix in pregnant rats still further, but had an inhibitory effect on the NO production in the uterus (myometrium). Treatment with the same dose of onapristone induced cervical ripening in rats.
These findings show that
treatment with a nitric oxide inhibitor such as L-NAME inhibits cervical ripening but stimulates uterine contractions in pregnant guinea pigs; PA1 the NO production, which has a relaxing effect on the myometrium, decreases in the pregnant uterus (myometrium) during the course of pregnancy, but it increases in the uterine cervix during normal and antiprogestin-induced parturition in rats; PA1 the divergent effects of a nitric oxide inhibitor such as L-NAME on uterine contractility and uterine cervix are due either to different actions of the NO-system or to the presence of different enzyme isoforms in the myometrium and cervix and PA1 the NO-system plays an important role in the control of uterine cervix, the NO production being increased during the ripening process of the cervix. PA1 (a) a local application of nitric oxide donors and/or substrates can be used to induce cervical ripening or PA1 (b) a local application of a nitric oxide-inhibitor can be used to prevent or inhibit cervical ripening, e.g. for the treatment of cervical insufficiency (too early cervical ripening) or preterm labour. PA1 (a) at least one nitric oxide donor and/or substrate for manufacture of a medicament to be administered locally (e. g. intracervically or intravaginally) for induction of cervical ripening or PA1 (b) at least one nitric oxide inhibitor for manufacture of a medicament to be administered locally (e. g. intracervically or intravaginally) for inhibition of cervical rirening for treatment of cervical insufficiency or preterm labor. PA1 onapristone (11.beta.-[4(Dimethylamino)phenyl]-17.alpha.-hydroxy-17.beta.-(3-hydroxypr opyl)-13.alpha.-estra-4,9-diene-3-one), RU 486 (11.beta.-[4-(Dimethylamino)phenyl]-17.beta.-hydroxy-17.alpha.-(1-propinyl )estra-4,9-diene-3-one), (Z)-11.beta.-[4-(Dimethylamino)phenyl]-17.beta.-hydroxy-17.alpha.-(3-hydro xy-1-propenyl)estr-4-ene-3-one (EP-A 0 404 283), 11.beta.-(4-Acetylphenyl)-17.beta.-hydroxy-17.alpha.-(1-propinyl)estra-4,9 diene-3-one (EP-A 0 190 759), 4',5'-Dihydro-11.beta.-[4-(dimethylamino)phenyl]-6.beta.-methylspiro[estra -4,9-diene-17.beta.,2'(3'H)-furan]-3-one, 4',5'-Dihydro-11.beta.-[4-(dimethylamino)phenyl]-7.beta.-methylspiro[estra -4,9-diene-17.beta.,2'(3'H)-furan]-3-one 11.beta.-(4-Acetylphenyl)-19,24-dinor-17,23-epoxy-17.alpha.-chola-4,9,20-t riene-3-one, (E)-11.beta.-[4-[[(Acetyloxy)imino]methyl]phenyl]-17.beta.-methoxy-17.alph a.-(methoxymethyl)estra-4,9-diene-3-one (E)-11.beta.-[4-[[[(Ethoxycarbonyl)oxy]imino]methyl]phenyl]-17.beta.-metho xy-17.alpha.-(methoxymethyl)estra-4,9-diene-3-one PA1 and a lot of other compounds having competitive progesterone antagonistic activity which are well known to the skilled persons. PA1 (A) induction of labour at term (time of ordinary birth, can be combined with sequential treatment with oxytocin or similar agents), PA1 (B) induction of labour in connection with a pathological pregnancy (e.g. fetal malformation); (preferred second trimester abortion), PA1 (C) induction of labour in connection with intrauterine fetal death, PA1 (D) induction of abortion (preferred first trimester abortion), PA1 (E) induction of preterm labour, PA1 (F) management of prolonged labour due to cervical dystocia, PA1 (G) induction of cervical ripening of a non-pregnant female or pregnant female to assist for surgical or diagnostic procedure, and PA1 (H) induction of cervical ripening for female to be treated by in vitro fertilisation. PA1 (I) treatment of cervical insufficiency (cervical incompetence) and PA1 (K) treatment of threatening preterm labour.
It is, therefore, concluded that:
Thus the present invention is directed to the use of nitric oxide donors and/or substrates or nitric oxide inhibitors for manufacture of a medicament for regulating cervical dilatation and/or extensibility.
The present invention involves the use of either
Suitable for the purposes of this invention as (a) nitric oxide donor and/or substrate or (b) nitric oxide inhibitor are all the compounds known to the persons skilled in the art as having the required properties; the compounds mentioned under "Description of related art" are preferred.
The nitric oxide donor and/or substrate (a) can further be used in combination with (i) at least one of an antiprogestin, a prostaglandin and/or a cytokine.
Examples for antiprogestins are
As prostaglandins suiprostone (PGE.sub.2) or gemeprost can be used for instance.
The most prominent representatives for the cytokines are the interleukin-8 or interleukin-1.beta..
These listings are not to be regarded as exhaustive.
The nitric oxide inhibitor (b) can further be used in combination with (ii) at least one of a progestin and/or a cyclooxygenase inhibitor (e. g. COX-1- and COX-2-inhibitors).
As a progestin, the naturally occuring progesterone is preferred but it is also possible to use one or more of the numerous synthetic progestins known to the artisan for use in oral contraceptives, for example levonorgestrel, cyproterone acetate, gestoden, drospirenone, desogestrel, 3-ketodesogestrel, dienogest etc.
The cyclooxygenase inhibitor can be for example aspirin (COX-1- and COX-2-inhibitor). Further examples of COX-2 inhibitors can be taken from: Robert Aslanian, Nicholas I. Carruthers, James J. Kaminski; Cyclooxygenase 2: A Novel Target for Therapeutic Intervention, Exp. Opin. Invest. Drugs, 1994, 3 (12), 1323-1325, for instance the compounds Sc-58125 (GD Searle), DuP-697, flusolide [6-(2,4-difuorophenoxy)-5-methylsulfonylamino-1-indanone], Sc-57666 (GD Searle), L-745 337 (Merck Frosst), NS-398 (Monsanto).
In its first aspect (a) the invention provides the use of these compounds of the invention in connection with birth or abortion. In this situation of pregnancy the cervix is pre-treated. The hormones during pregnancy alter the cervix which is then inclined to respond effectively to other stimulants.
Further the invention provides the use of the compounds under (a) of the invention in connection with surgical procedure and diagnostic procedure. Therefore, the compounds under (a) of the invention can, optionally in combination with the compounds mentioned under (i), be used for manufacture of medicaments for the following indications:
In its second aspect (b) the invention provides the use of these compounds of the invention in indications which require the prevention of too early cervical ripening or where the cervix has to be kept rigid. Therefore the compounds under (b) of this invention can, optionally in combination with the compounds mentioned under (ii), be used for manufacture of medicaments for the following indications:
In principle the compound can be used for human and non-human females. Human beings arc the preferred group for this treatment.
The nitric oxide donor and/or substrate (a) or the nitric oxide inhibitor (b) can be administered in any way in which, directly or indirectly, it will reach the cervix. Thus, it is conveniently applied intravaginally or directly to the cervix, e.g. typically as a gel or cream. It can also be injected into the cervical tissue or by a blunt needle into the cervical channel. It can also be applied extra-amniotically, i.e. between the uterine wall and the amniotic sac, using a catheter.
The preferred formulation is a gel or cream, but it can be applied as softenable capsules, liposomes or in a slow release formulation, or as an aqueous solution, e.g. a saline or protein-containing solution. The formulation can be achieved by methods known to the persons of ordinary skill in the art.
If a combined use of the compounds as mentioned above under (a) and (i) is intended the antiprogestin can be formulated for systemic or topical administration. The prostaglandin as well as the cytokine will be preferably formulated for local use.
If a combined use of the compounds as mentioned above under (b) and (ii) is intended, the compounds under (ii) can be formulated for local or systemic application.
The formulations of the compounds under (i) and (ii) can be achieved in ways known to those skilled in the art.
The compounds as under (a) of the invention exhibit pharmacological activity in induction of cervical ripening and may, therefore, be useful as a pharmaceutical agents.
The compounds as under (b) of the invention exhibit pharmacological activity in preventing cervical ripening and may, therefore, be useful as a pharmaceutical agents.
The measurement of the cervix ripening is described in Example 1.
Sodium nitroprusside and other nitric oxide donors show an effect on the cervical ripening at concentrations of from about 0.03 mM-100 mM (1.8 .mu.g-6 mg/application) when locally administered to pregnant female guinea-pigs. For these indications mentioned before under (A) to (F) the appropriate dosage will, of course, vary depending upon, for example, the compounds of the invention employed, the host, the mode of administration and the nature and severity of the condition being treated. However, in general, satisfactory results in animals are indicated to be obtained at a concentration from about 0.03-100 mM in the local formulation, preferred at a concentration of about 10 mM.
The compounds of the invention can be administered 6 to 48 hours before the final ripening of the cervix. The ripening can be proceeded by induction of labour with an oxytocic compound. The compounds can be administered in one or more dosages administered in a series with a distance of some hours or one day. The compounds of the invention may be administered by any conventional route, in particular in form of gel, ointment or local injection (at a concentration of about 0.03 to 100 mM).
Sodium nitroprusside and other nitric oxide donors show an effect on the cervical ripening at concentrations of from about 0.03 mM-100 mM (1.8 .mu.g-6 mmg/application) when administered to female guinea-pigs which are not pregnant. For these indications mentioned before under (G) to (H) the appropriate dosage will, of course, vary depending upon, for example, the compounds of the invention employed, the host, the mode of administration and the nature and severity of the condition being treated. However, in general, satisfactory results in animals are indicated to be obtained at a dosage from about 0.03 mM-100 mM (1.8 .mu.g-6 mmg/application), preferred at a concentration of about 10 mM. These compounds of the invention can be administered 24 or 48 hours before the final ripening of the cervix. The compounds can be administered in one or more dosages administered in a series with a distance of some hours or one day. The compound of the invention may be administered by any conventional route, in particular in form of gel, ointment or local injection.
Examples of human dosage ranges of typical NO-substrates and NO-donors are:
L-Arginine 500 mg-10 g/day Sodium nitroprusside range 500-2000 .mu.g/kg/day Nitroglycerine 0.5-10 mg/day Isosorbid mononitrate 10-100 mg/day Isosorbid dinitrate 10-100 mg/day.
Other NO-donors or NO-substrates are used in bioequivalent amounts.
Examples of dosage ranges of typical NO-inhibitors are:
 L-NAME 1 to 50 mg/kg/day L-NIO 1 to 50 mg/kg/day L-NA 1 to 50 mg/kg/day L-MMA 1 to 50 mg/kg/day L-NG 1 to 50 mg/kg/day L-NMA 1 to 50 mg/kg/day. Aminoguanidine 0.1 to 100 mg/kg/day.
Other NO-inhibitors are used in bioequivalent amounts.
Dosage range for RU 486: 25-600 mg/day per os.
Other antiprogestins are used in biologically equivalent amounts.
Dosage range for suiprostone: 100-1000 .mu.g/day i.m. or i.v.
Other prostaglandins such as PGE.sub.2 are used in bioequivalent amounts.
The prostaglandins can also be used locally intracervically in gel, intravaginally in gel or as tablets.
Dosage range for COX-2 inhibitors: 0.1-100 mg/kg/day.
Dosage range for IL-8: 100 ng-500 .mu.g/day; dosage range for IL-1.beta.: 100 ng-500 .mu.g/day.
Bioequivalent amounts of other compounds than those mentioned above can be determined by the methods as disclosed in the examples as such amounts which lead to comparable effects in the cervix as the specifically disclosed amounts under otherwise analogous conditions.